Purification method of n-(1(s)-ethoxycarbonyl-3-phenylpropyl)-l-alanine

ABSTRACT

The present invention is to provide a purification method of obtaining N-(1(S)-ethoxycarbonyl-3-phenylpropyl)-L-alanine of high quality in good yield with high productivity, which is accordingly suited for commercial scale application.  
     An impurity-contaminated N-(1(S)-ethoxycarbonyl-3-phenylpropyl)-L-alanine is crystallized from a mixed solvent of alcohol and water in a volume ratio of alcohol/water being 1 to 20 to remove a contaminating impurity into a mother liquor and give crystals of N-(1(S)-ethoxycarbonyl-3-phenylpropyl)-L-alanine.

FIELD OF THE INVENTION

[0001] The present invention relates to a purification method ofN-(1(S)-ethoxycarbonyl-3-phenylpropyl)-L-alanine represented by thefollowing formula (1).

[0002] N-(1(S)-ethoxycarbonyl-3-phenylpropyl)-L-alanine is a compound ofgreat use as an intermediate for the production of pharmaceuticals,particularly an intermediate for the production of severalantihypertensive drugs (angiotension converting enzyme inhibitors) suchas enalapril and ramipril.

PRIOR ART

[0003] The hithereto-known synthetic method ofN-(1(S)-ethoxycarbonyl-3-phenylpropyl)-L-alanine includes:

[0004] (a) the process involving Michael addition reaction of L-alanineto ethyl β-benzoylacrylate and subsequent catalytic reduction fortransformation of the carbonyl group of the benzoyl moiety to amethylene group (e.g. JP-A-03-22867)

[0005] (b) the process involving Michael addition reaction of L-alaninebenzyl ester to ethyl β-benzoylacrylate and subsequent catalyticreduction for concurrent transformation of the carbonyl group of thebenzoyl moiety to a methylene group and cleavage of the benzyl ester(e.g. JP-A-58-103364),

[0006] (c) the process which comprises reacting (S)-homophenylalanineethyl ester with (S)- or (RS)-propionic acid having a leaving group(halogen atom, sulfonyloxy group, or the like) in the α-position (e.g.JP-A-63-174956),

[0007] (d) the process which comprises reacting (S)-homophenylalanineethyl ester with (S)- or (RS)-propionic acid benzyl ester having aleaving group (halogen atom, sulfonyloxy group, or the like) in theα-position and then subjecting the reaction product to catalyticreduction for cleavage of the benzyl ester (e.g. JP-A-59-181247),

[0008] (e) the process which comprises reacting ethyl (R)- or(RS)-phenylbutyrate having a leaving group(halogen atom, sulfonyloxygroup, or the like) in the α-position with L-alanine benzyl or t-butylester of and subjecting the reaction product to catalytic reduction oracid treatment for cleavage of said benzyl or t-butyl ester (Chem.Pharm. Bull. 37(2), 280, 1989), and

[0009] (f) the process which comprises subjecting ethyl2-oxo-4-phenylbutyrate and L-alanine or L-alanine benzyl ester toreductive amination (e.g. JP-A-05-201882), etc.

[0010] In the above synthetic methods, production of the objectivecompound N-(1(S)-ethoxycarbonyl-3-phenylpropyl)-L-alanine is accompaniedby formation or remaining of various structurally analogous impuritiesas byproducts or residual contaminants.

[0011] As such impurities, there can be mentioned optical isomers,namely N-(1(R)-ethoxycarbonyl-3-phenylpropyl)-L-alanine represented bythe following formula (2),

[0012] N-(1(S)-ethoxycarbonyl-3-phenylpropyl)-D-alanine represented bythe following formula (3),

[0013] and N-(1(R)-ethoxycarbonyl-3-phenylpropyl)-D-alanine representedby the following formula (4);

[0014] the cyclohexyl derivative, namelyN-(1(S)-ethoxycarbonyl-3-cyclohexylpropyl)-L-alanine represented by thefollowing formula (5);

[0015] the carboxy derivative, namelyN-(1(S)--carboxy-3-phenylpropyl)-L-alanine represented by the followingformula (6);

[0016] the ester derivatives, namelyN-(1(S)-ethoxycarbonyl-3-phenylpropyl)-L-alanine ester represented bythe following formula (7),

[0017] in the formula, R represents an alkyl group or an aralkyl group;and

[0018] ethyl phenylbutyrate, etc.

[0019] The optical isomers are formed when the optical purity of thestarting material is low, when the stereoselectivity of the reaction isinsufficient, or due to racemization of the staring material orintermediate compound. Though it depends on the synthetic method,.generally N-(1(R) ethoxycarbonyl-3-phenylpropyl)-L-alanine of the aboveformula (2) and N-(1(S)-ethoxycarbonyl-3-phenylpropyl)-D-alanine of theabove formula (3) are formed as major byproducts.

[0020] The cyclohexyl derivative results from hydrogenation of thebenzene ring in catalytic reduction. The carboxy derivative results fromcleavage of an ester moiety in hydrolysis or catalytic reduction. Theester derivatives correspond to the compounds in which the terminalcarboxyl groups of the objective compounds are esterified. The formationof the derivatives result from remaining of the starting material owingto incomplete reaction or some side reaction. Referring to the formula(7), R represents an alkyl group of 1 to 8 carbon atoms (particularly analkyl group of 1 to 4 carbon atoms) such as ethyl, t-butyl or the likegroup or an aralkyl group of 7 to 10 carbon atoms such as benzyl or thelike group. The ethyl phenylbutyrate results from reduction of ethylβ-benzoylacrylate.

[0021] Of course, contamination of the product(N-(1(S)-ethoxycarbonyl-3-phenylpropyl)-L-alanine) with suchstructurally analogous impurities should be avoided as far as possibleand an effective purification technology is required for the purpose.

[0022] The hitherto-known purification method ofN-(1(S)-ethoxycarbonyl-3-phenylpropyl)-L-alanine includes, for example:

[0023] (1) the method of removing said optical isomers, particularly thediastereomer (1S/1R ratio=95/5→1S/1R ratio=99/1) by crystallization fromethyl acetate. (JP-A-03-22867)

[0024] (2) the method of removing said carboxy derivative bycrystallization from boiling water (AT402639B), etc.

[0025] Regarding the isolation method ofN-(1(S)-ethoxycarbonyl-3-phenylpropyl)-L-alanine, for exampleJP-A-05-201882 describes a method which comprises crystallizing anevaporation residue of the extracted organic phase from chilled ethanolor acetone, but there is no description referring to theimpurity-removing effect.

[0026] The present inventors made an investigation and as a result, theyrevealed that the above crystallization methods are not sufficient inimpurity-removing effect as reported in the past (AT402639B,JP-A-09-301938, etc.). Thus, for example, crystallization from ethylacetate may hardly remove said carboxy derivative, and crystallizationfrom water may hardly remove said cyclohexyl derivative, esterderivative, and ethyl phenylbutyrate which has low polarity.

[0027] Furthermore, in the conventional purification method, the lowsolubility of N-(1(S)-ethoxycarbonyl-3-phenylpropyl)-L-alanine in wateror a solvent such as ethanol makes it difficult to effectcrystallization in a high concentration range, thus having a problemfrom the standpoint of productivity on a commercial scale. In addition,the physical properties (liquid behaviors) of the resulting crystalslurry and the physical properties (powder characteristics) of theresulting crystal cannot necessarily be said to be satisfactory, andthere found problems such that the crystals cannot be easily dischargedfrom the crystallization tank, are not easy to dry and tend to formcakes in the drying stage, and that because of. small bulk specificgravity, containers of large capacity are required for packaging.

[0028] Thus, the conventional purification methods are not preferableenough from the standpoint of product purity, powder characteristics,yield, productivity and the like.

SUMMARY OF THE INVENTION

[0029] In view of the above state of art, the present invention has forits object to provide a purification method of obtainingN-(1(S)-ethoxycarbonyl-3-phenylpropyl)-L-alanine of high quality, namelyof high purity and having favorable powder characteristics, in goodyield with high productivity, which is accordingly suited for commercialscale application.

[0030] The present inventors made an intensive investigation for solvingthe above subject and as a result, found that carrying outcrystallization by using a mixed solvent of alcohol and water providesfor marked improvements in the solubility, impurity-removing effect,slurry behavior, and powder characteristics, all of which are parametersdeterminant of yield, quality, operability, and productivity. Thepresent invention has accordingly been developed.

[0031] The present invention, therefore, is directed to a purificationmethod of N-(1(S)-ethoxycarbonyl-3-phenylpropyl)-L-alanine representedby the formula (1)

[0032] which comprises crystallizing impurity-contaminatedN-(1(S)-ethoxycarbonyl-3-phenylpropyl)-L-alanine from a mixed solvent ofalcohol and water in a volume ratio of alcohol/water being 1 to 20 toremove a contaminating impurity into a mother liquor and give crystalsof N-(1(S)-ethoxycarbonyl-3-phenylpropyl)-L-alanine.

[0033] Moreover, the present invention is related to the abovepurification method wherein the contaminating impurity is at least onecompound selected from the group consisting ofN-(1(R)-ethoxycarbonyl-3-phenylpropyl)-L-alanine,N-(1(S)-ethoxycarbonyl-3-phenylpropyl)-D-alanine,N-(1(R)-ethoxycarbonyl-3-phenylpropyl)-D-alanine,N-(1(S)-ethoxycarbonyl-3-cyclohexylpropyl)-L-alanine,N-(1(S)-carboxy-3-phenylpropyl)-L-alanine,N-(1(S)-ethoxycarbonyl-3-phenylpropyl)-L-alanine ester, and ethylphenylbutyrate.

[0034] Further, the present invention is related to the abovepurification method wherein the crystallization is carried out underforced fluidity with a condition of not less than 0.1 kW/m³; the abovepurification method wherein the crystallization is carried out at atemperature of not lower than 20° C.; the above purification methodwherein the crystallization is carried out at a crystallizing speed ofnot more than 50% of a total crystal output/hour; the above purificationmethod wherein the crystallization is carried out at pH 3 to 6; theabove purification method wherein the crystallization is carried out byat least one of crystallization by cooling and crystallization byconcentration; the above purification method wherein the crystallizationis carried out by crystallization by cooling; and the above purificationmethod wherein the cooling speed for the crystallization by cooling isnot more than 40° C./hour.

[0035] Further, the present invention is related to the abovepurification method wherein the alcohol is a monohydric alcohol of 1 to8 carbon atoms; the above purification method wherein the alcohol is amonohydric alcohol of 1 to 4 carbon atoms; the above purification methodwherein the alcohol is ethanol; the above purification method whereinthe ethanol is denatured with a denaturing agent other than alcohol; thepurification method wherein a treatment with an adsorbent is carried outprior to the crystallization; and the above purification method whereinthe adsorbent is active charcoal.

DETAILED DESCRIPTION OF THE INVENTION

[0036] The present invention is now described in detail.

[0037] In the present invention, for obtainingN-(1(S)-ethoxycarbonyl-3-phenylpropyl)-L-alanine of high quality, namelyof high purity and having favorable powder characteristics, fromimpurity-contaminated N-(1(S)-ethoxycarbonyl-3-phenylpropyl)-L-alanine,in good yield with high productivity, the crystallization is carriedout. from a mixed solvent of alcohol and-water.

[0038] The above alcohol is not particularly restricted and includes,for example, monohydric alcohols of 1 to 12 carbon atoms such asmethanol, ethanol, 1-propanol, 2-propanol, 1-butanol, 2-butanol,isobutanol, t-butanol, 1-pentanol, 2-pentanol, 3-pentanol,2-methyl-1-butanol, isopentyl alcohol, t-pentyl alcohol,3-methyl-2-butanol, neopentyl alcohol, 1-hexanol, 2-methyl-1-pentanol,4-methyl-2-pentanol, 2-ethyl-1-butanol, 1-heptanol, 2-heptanol,3-heptanol, 1-octanol, 2-octanol, 2-ethyl-1-hexanol, 1-nonanol,3,3,5-trimethyl-1-hexanol, 1-decanol, 1-undecanol, 1-dodecanol, allylalcohol, propargyl alcohol, cyclohexanol, 1-methylcyclohexanol,2-methylcyclohexanol, 3-methylcyclohexanol, 4-methylcyclohexanol, benzylalcohol and so on.

[0039] Preferred are alcohols of 1 to 8 carbon atoms and include, forexample, methanol, ethanol, 1-propanol, 2-propanol, 1-butanol,2-butanol, isobutanol, t-butanol, 1-pentanol, 2-pentanol, 3-pentanol,2-methyl-1-butanol, isopentyl alcohol, t-pentyl alcohol,3-methyl-2-butanol, neopentyl alcohol, 1-hexanol, 2-methyl-1-pentanol,4-methyl-2-pentanol, 2-ethyl-1-butanol, 1-heptanol, 2-heptanol,3-heptanol, 1-octanol, 2-octanol, 2-ethyl-1-hexanol, allyl alcohol,propargyl alcohol, cyclohexanol, 1-methylcyclohexanol,2-methylcyclohexanol, 3-methylcyclohexanol, 4-methylcyclohexanol, benzylalcohol, and so on.

[0040] From the standpoint of product quality, yield, and productivity,a monohydric alcohol of 1 to 6 carbon atoms is more preferred and therecan be mentioned, for example, methanol, ethanol, 1-propanol,2-propanol, 1-butanol, 2-butanol, isobutanol, t-butanol, 1-pentanol,2-pentanol, 3-pentanol, 2-methyl-1-butanol, isopentyl alcohol, t-pentylalcohol, 3-methyl-2-butanol, neopentyl alcohol, 1-hexanol,2-methyl-1-pentanol, 4-methyl-2-pentanol, 2-ethyl-1-butanol, allylalcohol, propargyl alcohol, cylohexanol, and so on.

[0041] The monohydric alcohol of 1 to 4 carbon atoms is still morepreferred in that it can be appropriately heated for enhancedsolubility, that both removal of the solvent from wet crystals andrecovery of the solvent from the crystallization filtrate can be easilyaccomplished, that it is hardly solidified on cooling to a temperaturebelow room temperature, that it is easy to work with because of its lowviscosity, and that it is advantageous in terms of solvent cost andavailability. For example, there can be mentioned methanol, ethanol,1-propanol, 2-propanol, 1-butanol, 2-butanol, isobutanol, t-butanol,allyl alcohol, propargyl alcohol, and so on.

[0042] When any alcohol other than ethanol is used, depending onconditions, byproducts due to transesterification ofN-(1(S)-ethoxycarbonyl-3-phenylpropyl)-L-alanine, which are hardlyremoved, tend to form (e.g.N-(1(S)-methoxycarbonyl-3-phenylpropyl)-L-alanine andN-(1(S)-methoxycarbonyl-3-phenylpropyl)-L-alanine methyl ester).Therefore, it is most preferable to use ethanol.

[0043] When ethanol is used, that ethanol may be denatured with adenaturing agent. Usable as the denaturing agent include isopropylalcohol, methanol, ethyl acetate, methyl isobutyl ketone, aliphatichydrocarbons (e.g. hexane and heptane), and aromatic hydrocarbons (e.g.toluene and benzene), and so forth. Among these, it is preferable to usea denaturing agent other than alcohols. More preferred are aliphatichydrocarbons and aromatic hydrocarbons, with toluene being particularlypreferred. The level of addition of the denaturing agent is generallynot higher than 10% relative to the volume of ethanol.

[0044] In the present invention, as an auxiliary solvent, water is usedin combination with said alcohol. Concomitant use of water increases thesolubility of N-(1(S)-ethoxycarbonyl-3-phenylpropyl)-L-alanine to anappropriate level and leads to improvements not only in yield andproductivity but also in impurity-removing effect, slurry behavior, andphysical properties of crystals (powder characteristics).

[0045] The volume ratio of alcohol to water for crystallization dependson the kind of alcohol to be used but it is necessary that thealcohol/water volume ratio is 1 to 20. The upper limit is preferably 18,more preferably 16, still more preferably 14, particularly preferably10. From quality points of view, it is more preferably 5, still morepreferably 4, particularly preferably 3. The lower limit is preferably1.5, more preferably 2, from quality points of view. The preferablerange is 1.5 to 10, more preferably 2 to 5, still more preferably 2 to4, particularly preferably 2 to 3 by which condition the crystallizationcan suitably be carried out. It is preferred that the ratio is selectedso as to attain the yield of not less than about 70%, preferably notless than 80%, more preferably not less than 90%.

[0046] In the present invention, the crystallization ofN-(1(S)-ethoxycarbonyl-3-phenylpropyl)-L-alanine is preferably carriedout at pH 3 to 6, more preferably at pH 4 to 5 of the solution composedof the crystals and the above mixed solvent, from the standpoint ofyield and quality (inclusive of inhibition of formation of byproductimpurities). When the pH of the solution is too low or too high owing tothe presence of impurities and so on, the pH can be adjusted, forexample, with an acid, such as hydrochloric acid or sulfuric acid, or analkali, e.g. an alkali metal hydroxide, such as sodium hydroxide orlithium hydroxide.

[0047] In the present invention, the crystallization ofN-(1(S)-ethoxycarbonyl-3-phenylpropyl)-L-alanine is preferably carriedout under forced fluidity. From quality points of view, the fluidity interms of agitation power per unit volume is preferably not less thanabout 0.1 kW/m³, more preferably not less than about 0.2 kW/m³, stillmore preferably not less than about 0.3 kW/m³. The upper limit is notparticularly restricted, but-is preferably not higher than about 20kW/m³, more preferably not higher than about 10 kW/m³. The forcedfluidity mentioned above is generally established by rotation of astirring impeller, but it is not always necessary to employ the stirringimpeller provided that the above fluidity is obtained. For example, asystem utilizing circulation of the solution can be exploited.

[0048] From quality (product purity, powder characteristics) points ofview, the crystallization ofN-(1(S)-ethoxycarbonyl-3-phenylpropyl)-L-alanine is preferably carriedout under warming condition and is preferably carried out at atemperature of not lower than about 20° C., more preferably not lowerthan about 30° C. The upper limit is preferably not higher than about80° C, more preferably not higher than about 70° C. The crystallizationcan suitably be carried out at about 20° to 80° C.

[0049] The crystallization according to the present invention can becarried out by the routine crystallizing technique, that is to say atleast any one of such techniques as-crystallization by cooling,crystallization by neutralization, and crystallization by concentration(inclusive of crystallization by solvent exchange). It is preferable touse at least one of crystallization by cooling and crystallization byconcentration, and is particularly preferable to use crystallization bycooling.

[0050] To maximize the effect of the invention, it is preferable thatthe contamination of various impurities into crystals ofN-(1(S)-ethoxycarbonyl-3-phenylpropyl)-L-alanine be minimized bycontrolling the crystallizing speed, that is to say the crystal outputper unit period. The crystallizing speed is preferably not more thanabout 50% of the total crystal output/hour, more preferably not morethan about 25% of the total crystal output/hour. The lower limit ispreferably 1% of the total crystal output/hour, more preferably 2% ofthe total crystal output/hour.

[0051] In the case of crystallization by cooling, the cooling speed ispreferably not more than about 40° C./hour, more preferably not morethan about 20° C./hour, still more preferably not more than about 10°C./hour, particularly preferably not more than 5° C./hour, from qualitypoints of view. The lower limit is preferably not less than about 1°C./hour, more preferably not less than about 2° C./hour. In this case,since abrupt crystallization with collapse of a large built-up ofsupersaturation is undesirable from quality points of view, it is goodpractice to add seed crystals to provide for smooth nucleation wherenecessary.

[0052] The crystal concentration at completion of crystallization is notparticularly restricted and this is also dependent on the kind ofalcohol to be used but the weight ratio ofN-(1(S)-ethoxycarbonyl-3-phenylpropyl)-L-alanine relative to the volumeof the solvent is preferably about 5 to 40 w/v %, more preferably about10 to 35 w/v %, still more preferably 20 to 30 w/v %.

[0053] The purification method of the present invention provides for ahigh impurity-removing effect and is particularly effective in removingoptical isomers (N-(1(R)-ethoxycarbonyl-3-phenylpropyl)-L-alanine,N-(1(S)-ethoxycarbonyl-3-phenylpropyl)-D-alanine andN-(1(R)-ethoxycarbonyl-3-phenylpropyl)-D-alanine), cyclohexyl derivative(N-(1(S)-ethoxycarbonyl-3-cyclohexylpropyl)-L-alanine), carboxyderivative (N-(1(S)-carboxy-3-phenylpropyl)-L-alanine), ester derivative(N-(1(S)-ethoxycarbonyl-3-phenylpropyl)-L-alanine ester), and ethylphenylbutyrate. In particular, it is very effective in removing thecyclohexyl derivative which is otherwise extremely difficult to remove.Moreover, the technology is effective in removing iron and otherinorganic contaminants as well.

[0054] To assist in impurity removal, it is effective to treat thesubstrate with an adsorbent, preferably with active charcoal, prior tothe crystallization.

[0055] The crystals of N-(1(S)-ethoxycarbonyl-3-phenylpropyl)-L-alanineobtainable by the purification method of the invention can be obtainedas wet crystals by the conventional solid-liquid separation/cake washingprocedure (centrifugation, pressure filtration, suction filtration,etc.) and can also be obtained as dry crystals by subjecting the wetcrystals further to the conventional drying procedure (e.g. air drying,drying under reduced pressure, drying in vacuo, etc.). In conducting thesolid-liquid separation, the yield can be maximized by cooling thesystem to a temperature not higher than about 20° C., preferably 0 to10° C.

[0056] Though not particularly restricted, the purification method ofthe invention can suitably be used as an isolation method or arecrystallization method for obtaining the crystals ofN-(1(S)-ethoxycarbonyl-3-phenylpropyl)-L-alanine synthesized by anyknown production method mentioned above, particularly ofN-(1(S)-ethoxycarbonyl-3-phenylpropyl)-L-alanine synthesized by themethod involving Michael addition reaction as mentioned hereinbeforeunder (a) or (b) in Prior Art.

[0057] It is considered that the effect of the present invention resultsfrom the fact that water content inside the crystals is high whenN-(1(S)-ethoxycarbonyl-3-phenylpropyl)-L-alanine is crystallized fromthe mixed solvent of alcohol and water comparing to when crystallizedfrom alcohol.

BEST MODE FOR CARRYING OUT THE INVENTION

[0058] The following examples illustrate the present invention infurther detail without defining the scope of the invention.

[0059] (Production Example) Production ofN-(1-ethoxycarbonyl-3-phenylpropyl)-L-alanine

[0060] To a solution of 25.9 g of ethyl trans-β-benzoylacrylate in 770ml of ethanol was added a solution of 6.03 g of L-alanine lithium saltin 426 ml of ethanol over 30 minutes at room temperature. Aftercompletion of addition, the mixture was stirred for an additional 5minutes, and then 5.29 ml of concentrated hydrochloric acid was added,followed by cooling with ice-water. As seed crystals, 679 mg ofN-(1(S)-ethoxycarbonyl-3-oxo-3-phenylpropyl)-L-alanine was added, andthe mixture was stirred for 4 hours. The crystals separating out werecollected by filtration, washed with ethanol, and dried, whereby 12.7 gof N-(1-ethoxycarbonyl-3-oxo-3-phenylpropyl)-L-alanine (1S/1Rratio=95/5) was obtained.

[0061] In 110 ml of 1% (v/v) sulfuric acid-ethanol was dissolved 2.0 gof thus-obtained N-(1-ethoxycarbonyl-3-oxo-3-phenylpropyl)-L-alanine(1S/1R ratio=95/5), followed by addition of 0.5 g of 10% Pd/C, andcatalytic reduction was carried out at room temperature underatmospheric pressure. After the reaction, the catalyst was removed bysuction filtration, the filtrate was washed with ethanol and theobtained solution was concentrated. The concentrate was neutralized byadding water and sodium hydroxide and the crystals separating out werecollected by filtration, washed with water, and dried to give 1.5 g ofN-(1-ethoxycarbonyl-3-phenylpropyl)-L-alanine (1S/1R ratio=99/1). Themean particle diameter (D_(P) ⁵⁰) of the crystals was 30 μm, the loosepacking bulk specific gravity was about 0.3, and the flowability of thecrystals was not satisfactory.

EXAMPLE 1

[0062] Five grams of N-(1(S)-ethoxycarbonyl-3-phenylpropyl)-L-alanine(purity 96.7%; impurities contained:N-(1(R)-ethoxycarbonyl-3-phenylpropyl)-L-alanine 0.8%,N-(1(S)-ethoxycarbonyl-3-cyclohexylpropyl)-L-alanine 0.84%,N-(1(S)-carboxy-3-phenylpropyl)-L-alanine 0.2%,N-(1(S)-ethoxycarbonyl-3-phenylpropyl)-L-alanine ethyl ester 0.5%, andethyl phenylbutyrate 0.15%) was dissolved in 20 ml of a mixed solvent ofethanol and water (ethanol/water volume ratio 7) under warming (about65° C.). The solution was cooled to 20° C. over 2 hours under stirringfor crystallization (pH during crystallization 4 to 5). Then, understirring, the system was further cooled to 10° C. and the resultingcrystals were collected by filtration, washed with cold mixed solvent ofethanol and water (ethanol/water volume ratio 7), and dried in vacuo (40to 60° C., overnight), whereupon dry crystals ofN-(1(S)-ethoxycarbonyl-3-phenylpropyl)-L-alanine were obtained. Yield84%, purity 100.0%. None ofN-(1(R)-ethoxycarbonyl-3-phenylpropyl)-L-alanine,N-(1(S)-carboxy-3-phenylpropyl)-L-alanine,N-(1(S)-ethoxycarbonyl-3-phenylpropyl)-L-alanine ethyl ester, and ethylphenylbutyrate were detected, and theN-(1(S)-ethoxycarbonyl-3-cyclohexylpropyl)-L-alanine content was 0.40%(removal rate 52%). The mean particle diameter (D_(P) ⁵⁰) of thecrystals was 170 μm, the loose packing bulk specific gravity was about0.5, and the flowability of crystals was satisfactory.

EXAMPLE 2

[0063] Five grams of the sameN-(1(S)-ethoxycarbonyl-3-phenylpropyl)-L-alanine as used in Example 1was dissolved in 28 ml of a mixed solvent of isobutanol and water(isobutanol/water volume ratio 10) under warming (about 65° C.). Thesolution was cooled to 20° C. over 2 hours under stirring forcrystallization (pH during crystallization 4 to 5). The crystals werecollected by filtration, washed with a cold mixed solvent of isobutanoland water (isobutanol/water volume ratio 10), and dried in vacuo (40 to60° C., overnight) to give dry crystals ofN-(1(S)-ethoxycarbonyl-3-phenylpropyl)-L-alanine. Yield 81%, purity99.9%. None of N-(1(R)-ethoxycarbonyl-3-phenylpropyl)-L-alanine,N-(1(S)-carboxy-3-phenylpropyl)-L-alanine,N-(1(S)-ethoxycarbonyl-3-phenylpropyl)-L-alanine ethyl ester, and ethylphenylbutyrate were detected, and theN-(1(S)-ethoxycarbonyl-3-cyclohexylpropyl)-L-alanine content was 0.27%(removal rate 68%). The mean particle diameter (D_(P) ⁵⁰) of crystalswas 130 μm, the loose packing bulk specific gravity was about 0.5, andthe flowability of crystals was satisfactory.

EXAMPLE 3

[0064] Five grams of the sameN-(1(S)-ethoxycarbonyl-3-phenylpropyl)-L-alanine as used in Example 1was dissolved in 20 ml of a mixed solvent of 1-propanol and water(1-propanol/water volume ratio 10) under warming (about 65° C.). Thesolution was cooled to 20° C. over 2 hours under stirring forcrystallization (pH during crystallization 4 to 5). The crystals werecollected by filtration, washed with a cold mixed solvent of 1-propanoland water (1-propanol/water volume ratio 10), and dried in vacuo (40 to60° C., overnight) to give dry crystals ofN-(1(S)-ethoxycarbonyl-3-phenylpropyl)-L-alanine. Yield 85%, purity99.7%. None of N-(1-(R)-ethoxycarbonyl-3-phenylpropyl)-L-alanine,N-(1(S)-carboxy-3-phenylpropyl)-L-alanine,N-(1(S)-ethoxycarbonyl-3-phenylpropyl)-L-alanine ethyl ester, and ethylphenylbutyrate were detected, and theN-(1(S)-ethoxycarbonyl-3-cyclohexylpropyl)-L-alanine content was 0.38%(removal rate 55%). The mean particle diameter (D_(P) ⁵⁰) of thecrystals was 120 μm, the loose packing bulk specific gravity was about0.5, and the flowability of crystals was satisfactory.

Comparative Example 1

[0065] Five grams of N-(1(S)-ethoxycarbonyl-3-phenylpropyl)-L-alanine(as an impurity, 0.84% ofN-(1(S)-ethoxycarbonyl-3-cyclohexylpropyl)-L-alanine was contained) wasdissolved in 50 ml of isobutanol under warming (about 65° C). When thesolution was cooled to 20° C. over 2 hours under stirring forcrystallization, it solidified to form cakes. The crystals werecollected by filtration, washed with cold isobutanol, and dried in vacuo(40 to 60° C., overnight) to give dry crystals ofN-(1(S)-ethoxycarbonyl-3-phenylpropyl)-L-alanine. Yield 63%, theN-(1(S)-ethoxycarbonyl-3-cyclohexylpropyl)-L-alanine content was 0.40%(removal rate 52%).

Comparative Example 2

[0066] Five grams of N-(1(S)-ethoxycarbonyl-3-phenylpropyl)-L-alanine(as an impurity, 0.84% ofN-(1(S)-ethoxycarbonyl-3-cyclohexylpropyl)-L-alanine was contained) wasdissolved in 30 ml of ethanol under warming (about 65° C.). When thesolution was cooled to 20° C. over 2 hours under stirring forcrystallization, it solidified to form cakes. The crystals werecollected by filtration, washed with cold ethanol, and dried in vacuo(40 to 60° C., overnight) to give dry crystals ofN-(1(S)-ethoxycarbonyl-3-phenylpropyl)-L-alanine. Yield 67%, theN-(1(S)-ethoxycarbonyl-3-cyclohexylpropyl)-L-alanine content was 0.47%(removal rate 44%).

Comparative Example 3

[0067] Five grams of N-(1(S)-ethoxycarbonyl-3-phenylpropyl)-L-alanine(as an impurity, 0.84% ofN-(1(S)-ethoxycarbonyl-3-cyclohexylpropyl)-L-alanine was contained) wasdissolved in 32 ml of a mixed solvent of ethanol and cyclohexane(ethanol/cyclohexane volume ratio 2) under warming (about 65° C). Thesolution was cooled to 20° C. over 2 hours under stirring forcrystallization. The crystals were collected by filtration, washed witha cold mixed solvent of ethanol and cyclohexane (ethanol/cyclohexanevolume ratio 2), and dried in vacuo (40 to 60° C., overnight) to givedry crystals of N-(1(S)-ethoxycarbonyl-3-phenylpropyl)-L-alanine. Yield70%. The N-(1(S)-ethoxycarbonyl-3-cyclohexylpropyl)-L-alanine contentwas 0.48% (removal rate 43%).

EXAMPLE 4

[0068] Thirty grams of N-(1(S)-ethoxycarbonyl-3-phenylpropyl)-L-alanine(purity 96.4%; impurities contained:N-(1(R)-ethoxycarbonyl-3-phenylpropyl)-L-alanine 0.10% andN-(1(S)-ethoxycarbonyl-3-cyclohexylpropyl)-L-alanine 0.11%) wasdissolved in 100 ml of a mixed solvent of ethanol and water(ethanol/water volume ratio 2.96) under warming (about 65° C.). Thesolution was treated with 3 g of 50% hydrous active charcoal for 1 hour,the mixture was then filtered when hot, and washed with 10 ml of a mixedsolvent of ethanol and water (ethanol/water volume ratio 16.9). Theresulting solution was rapidly cooled to 20° C. under vigorous stirring(0.3 kW/m³) (cooling speed 40° C./hr) and further stirred for 2 hours(pH during crystallization 4 to 5). The crystals were collected byfiltration, washed with a cold mixed solvent of ethanol and water(ethanol/water volume ratio 16.9), and dried in vacuo to give drycrystals of N-(1(S)-ethoxycarbonyl-3-phenylpropyl)-L-alanine. Yield 85%;N-(1(R)-ethoxycarbonyl-3-phenylpropyl)-L-alanine was not detected andthe N-(1(S)-ethoxycarbonyl-3-cyclohexylpropyl)-L-alanine content was0.050% (removal rate 55%).

EXAMPLE 5

[0069] The procedure of Example 4 was repeated except that thecrystallization was carried out at a cooling speed of 10° C./hour. Yield85%; N-(1(R)-ethoxycarbonyl-3-phenylpropyl)-L-alanine was not detectedand the N-(1(S)-ethoxycarbonyl-3-cyclohexylpropyl)-L-alanine content was0.044% (removal rate 60%).

EXAMPLE 6

[0070] The procedure of Example 4 was repeated except that thecrystallization was carried out at a cooling speed of 5° C./hour. Yield85%; N-(1(R)-ethoxycarbonyl-3-phenylpropyl)-L-alanine was not detectedand the N-(1(S)-ethoxycarbonyl-3-cyclohexylpropyl)-L-alanine content was0.035% (removal rate 68%).

EXAMPLE 7

[0071] Five grams of N-(1(S)-ethoxycarbonyl-3-phenylpropyl)-L-alanine(purity 96.7%; impurities contained:N-(1(R)-ethoxycarbonyl-3-phenylpropyl)-L-alanine 0.8%,N-(1(S)-ethoxycarbonyl-3-cyclohexylpropyl)-L-alanine 0.84%,N-(1(S)-carboxy-3-phenylpropyl)-L-alanine 0.2%,N-(1(S)-ethoxycarbonyl-3-phenylpropyl)-L-alanine ethyl ester 0.5%, andethyl phenylbutyrate 0.15%) was dissolved in 17 ml of a mixed solvent ofethanol and water (ethanol/water volume ratio 2.3) under warming (about65° C.). This solution was treated with 1 g of 50% hydrous activecharcoal for 10 minutes, the mixture was then filtered when hot, andwashed with 3 ml of the mixed solvent of ethanol and water(ethanol/water volume ratio 2.3). The filtrate thus obtained was cooledto 20° C. over 2 hours under stirring (0.2 kW/m³) for crystallization(pH during crystallization 4 to 5). The slurry was further cooled to 10°C. under stirring (0.2 kW/m³), after which the crystals were collectedby filtration, washed with a cold mixed solvent of ethanol and water(ethanol/water volume ratio 16.9), and dried in vacuo (40 to 60° C.,overnight) to give dry crystals ofN-(1(S)-ethoxycarbonyl-3-phenylpropyl)-L-alanine. Yield 83%, purity99.9%. None of N-(1(R)-ethoxycarbonyl-3-phenylpropyl)-L-alanine,N-(1(S)-carboxy-3-phenylpropyl)-L-alanine,N-(1(S)-ethoxycarbonyl-3-phenylpropyl)-L-alanine ethyl ester, and ethylphenylbutyrate were detected, theN-(1(S)-ethoxycarbonyl-3-cyclohexylpropyl)-L-alanine content was 0.40%(removal rate 52%), and the iron content was 0.5 ppm. The mean particlediameter (D_(P) ⁵⁰) of the crystals was 160 μm, the loose packing bulkspecific gravity was about 0.5, and the flowability of crystals wassatisfactory.

Comparative Example 4

[0072] Five grams of the sameN-(1(S)-ethoxycarbonyl-3-phenylpropyl)-L-alanine as used in Example 7was added to 55 ml of water and dissolved by adding 1.9 ml ofconcentrated hydrochloric acid. This solution was treated with 1 g of50% hydrous active charcoal for 10 minutes, and the mixture was thenfiltered and washed with 5 ml of water. To this filtrate was added 1 mlof 30% aqueous solution of sodium hydroxide over 1 hour at 25 to 28° C.under stirring (0.2 kW/m³) to adjust the pH of the solution to 4.7. Thesolution was stirred at 22° C. for 1 hour, and the crystals werecollected by filtration, washed with 5 ml of water, and dried in vacuo(40 to 60° C., overnight) to give dry crystals ofN-(1(S)-ethoxycarbonyl-3-phenylpropyl)-L-alanine. Yield 86%, purity99.1%; neither N-(1(S)-carboxy-3-phenylpropyl)-L-alanine nor ethylphenylbutyrate was detected. TheN-(1(R)-ethoxycarbonyl-3-phenylpropyl)-L-alanine content was 0.1%, theN-(1(S)-ethoxycarbonyl-3-phenylpropyl)-L-alanine ethyl ester content was0.2%, the N-(1(S)-ethoxycarbonyl-3-cyclohexylpropyl)-L-alanine contentwas 0.84% (removal rate 0%), and the iron content was 30 ppm. The meanparticle diameter (D_(P) ⁵⁰) of the crystals was 70 μm, the loosepacking bulk specific gravity was about 0.3, and the flowability ofcrystals was not as good as desired.

[0073] In the purification method according to each of Examples as shownabove, N-(1(S)-ethoxycarbonyl-3-phenylpropyl)-L-alanine of high quality,namely of high purity having favorable powder characteristics(especially, the mean particle diameter was within a favorable range of100 to 1000 μm, the loose packing bulk specific gravity was within afavorable range of 0.4 to 0.7 and the flowability of crystals wassatisfactory) could be purified and obtained in good yield with highproductivity.

Reference Example 1

[0074] The powder characteristics of the crystals obtained in the samemanner as in Example 7 were studied using Hosokawa Micron's powdertester. The results are shown below.

[0075] Apparent specific gravity, loose: 0.47

[0076] Apparent specific gravity, packed: 0.55

[0077] Degree of compaction, %: 15

[0078] Degree of compaction, index: 20

[0079] Angle of repose, degree: 43

[0080] Angle of repose, index: 16

[0081] Spatula angle, degree: 46

[0082] Spatula angle, index: 17

[0083] Degree of uniformity, unit: 2.1

[0084] Degree of uniformity, index: 23

[0085] Flowability index: 76

[0086] Degree of flowability: fairly good

[0087] It was found from the above results that the powdercharacteristics of the above crystals were excellent.

Reference Example 2

[0088] The powder characteristics of the crystals obtained in the samemanner as in Comparative Example 4 were studied using Hosokawa Micron'spowder tester. The results are shown below.

[0089] Apparent specific gravity, loose: 0.24

[0090] Apparent specific gravity, packed: 0.39

[0091] Degree of compaction, %: 39

[0092] Degree of compaction, index: 2

[0093] Angle of repose, degree: 50

[0094] Angle of repose, index: 12

[0095] Spatula angle, degree: 66

[0096] Spatula angle, index: 12

[0097] Degree of uniformity, unit: 1.6

[0098] Degree of uniformity, index: 24

[0099] Flowability index: 50

[0100] Degree of flowability: not as good as desired

[0101] It was found from the results shown above that the powdercharacteristics of the above crystals were inferior comparing toReference Example 1.

Reference Example 3

[0102] One-hundred milliliters each of mixed solvents of ethanol andwater in predetermined volume ratios were respectively adjusted topredetermined temperatures and each solvent was added until a pureproduct of N-(1(S)-ethoxycarbonyl-3-phenylpropyl)-L-alanine was nolonger dissolved. After 30 minutes of standing, the supernatant wastaken and the solubility (weight %) was determined according to theweight after concentration to dryness/the weight of the solution. Theresults are shown in Table 1. TABLE 1 Ethanol/water volume ratio andsolubility (weight %) Volume ratio 0.43 1.00 2.33 4.00 5.67 15.7 249Solubility 65° C. 9 21 42 45 42 26 17 20° C. — 3 4 5 5 4 3 10° C. 1 2 34 3 3 2

[0103] As shown in the above, when ethanol/water volume ratio is 1 to20, the solubility is highly dependent on the temperature, henceimproved yield and productivity (crystallization concentration) areexpected by using the mixed solvent as the crystallization solvent.

Industrial Applicability

[0104] By the purification method of the invention,N-(1(S)-ethoxycarbonyl-3-phenylpropyl)-L-alanine of high quality, namelyof high purity and having favorable powder characteristics, can bepurified and obtained in good yield with high productivity.

1. A purification method ofN-(1(S)-ethoxycarbonyl-3-phenylpropyl)-L-alanine represented by theformula (1)

which comprises crystallizing impurity-contaminatedN-(1(S)-ethoxycarbonyl-3-phenylpropyl)-L-alanine from a mixed solvent ofalcohol and water in a volume ratio of alcohol/water being 1 to 20 toremove a contaminating impurity into a mother liquor and give crystalsof N-(1(S)-ethoxycarbonyl-3-phenylpropyl)-L-alanine.
 2. The purificationmethod according to claim 1 wherein the contaminating impurity is atleast one compound selected from the group consisting ofN-(1(R)-ethoxycarbonyl-3-phenylpropyl)-L-alanine represented by thefollowing formula (2),

N-(1(S)-ethoxycarbonyl-3-phenylpropyl)-D-alanine represented by thefollowing formula (3),

N-(1(R)-ethoxycarbonyl-3-phenylpropyl)-D-alanine represented by thefollowing formula (4),

N-(1(S)-ethoxycarbonyl-3-cyclohexylpropyl)-L-alanine represented by thefollowing formula (5),

N-(1(S)-carboxy-3-phenylpropyl)-L-alanine represented by the followingformula (6),

N-(1(S)-ethoxycarbonyl-3-phenylpropyl)-L-alanine ester of the followinggeneral formula (7)

in the formula, R represents an alkyl group or an aralkyl group, andethyl phenylbutyrate.
 3. The purification method according to claim 1 or2 wherein the crystallization is carried out under forced fluidity witha condition of not less than 0.1 kW/m³.
 4. The purification methodaccording to any of claims 1 to 3 wherein the crystallization is carriedout at a temperature of not lower than 20° C.
 5. The purification methodaccording to any of claims 1 to 4 wherein the crystallization is carriedout at a crystallizing speed of not more than 50% of a total crystaloutput/hour.
 6. The purification method according to any of claims 1 to5 wherein the crystallization is carried out at pH 3 to
 6. 7. Thepurification method according to any of claims 1 to 6 wherein thecrystallization is carried out by at least one of crystallization bycooling and crystallization by concentration.
 8. The purification methodaccording to claim 7 wherein the crystallization is carried out bycrystallization by cooling.
 9. The purification method according toclaim 8 wherein the cooling speed for the crystallization by cooling isnot more than 40° C./hour.
 10. The purification method according to anyof claims 1 to 9 wherein the alcohol is a monohydric alcohol of 1 to 8carbon atoms.
 11. The purification method according to claim 10 whereinthe alcohol is a monohydric alcohol of 1 to 4 carbon atoms.
 12. Thepurification method according to claim 11 wherein the alcohol isethanol.
 13. The purification method according to claim 12 wherein theethanol is denatured with a denaturing agent other than alcohol.
 14. Thepurification method according to any of claims 1 to 13 wherein atreatment with an adsorbent is carried out prior to the crystallization.15. The purification method according to claim 14 wherein the adsorbentis active charcoal.